1. Field of the Invention
This invention relates to a novel chemical synthesis of stable, water-soluble chemiluminescent 1,2-dioxetanes and to novel intermediates obtained in the course of synthesizing such 1,2-dioxetanes.
2. Description of Related Art
1,2-Dioxetanes, cyclic organic peroxides whose central structure is a four-membered ring containing pairs of contiguous carbon and oxygen atoms (the latter forming a peroxide linkage), are a known, but until recently seldom utilized, class of compounds. Some 1,2-dioxetanes can be made to exhibit chemiluminescent decomposition, e.g., by the action of enzymes, as described in the following copending, commonly-assigned U.S. patent applications: Bronstein, Ser. No. 889,823, "Method of Detecting a Substance Using Enzymatically-Induced Decomposition of Dioxetanes", filed Jul. 24, 1986; Bronstein et al., Ser. No. 140,035, "Dioxetanes for Use in Assays", filed Dec. 31, 1987 now abandoned; Edwards, Ser. No. 140,197, "Synthesis of 1,2-Dioxetanes and Intermediates Therefor", filed Dec. 31, 1987 now abandoned; Edwards, et al., Ser. No. 213,672, "Novel Chemiluminescent Fused Polycyclic Ring-Containing 1,2-dioxetanes and Assays in Which They Are Used", filed Jun. 30, 1988, now U.S. Pat. No. 4,952,707; as well as in Bronstein, I.Y. et al., "Novel Enzyme Substrates and Their Application in Immunoassay", J. Biolum. Chem.. 2:186 (1988).
The amount of light emitted during such chemiluminescence is a measure of the concentration of a luminescent substance which, in turn, is a measure of the concentration of its precursor 1,2-dioxetane. Thus, by measuring the intensity of luminescence, the concentration of the 1,2-dioxetane, and hence the concentration of a substance being assayed (e.g., a biological species bound to the 1,2-dioxetane member of a specific binding pair in a bioassay) can be determined. The appropriate choice of substituents on the 1,2-dioxetane ring allows, inter alia, for adjustment of the chemical stability of the molecule which, in turn, affords a means of controlling the onset of chemiluminescence, thereby enhancing the usefulness of such chemiluminescence for practical purposes, e.g., immunoassays, nucleic acid probe assays, enzyme assays, and the like.
The preparation of 1,2-dioxetanes by photo-oxidation of olefinic double bonds is known. Mazur, S. et al., J. Am. Chem. Soc., 92:3225 (1970). However a need exists for a facile, general synthesis of substituted 1,2-dioxetanes from olefinically-unsaturated precursors derived from readily available or obtainable starting materials through tractable intermediates. In this connection, a particular need exists for a commercially useful method for producing 1,2-dioxetanes of the general formula: ##STR4## wherein T, R.sup.3, Y and Z are defined herein below, from enol ether-type precursors of the general formula: ##STR5##
McMurry et al. [McMurry, J.E., et al., J. Org. Chem., 43:3255 (1978)] described titanium-induced reductive coupling of carbonyl groups to form olefins. Schaap, A.P., EPO 254,051, published Jan. 27, 1988, and Bronstein, I.Y., 1986, disclose the use of this reaction to produce compounds of formula (II) by the following general reaction: ##STR6##
Several problems with aforementioned unsymmetrical McMurry coupling are especially important in the radical based mechanism which operates in the above equation when compared with similar mixed couplings between aliphatic and diaryl ketones where the mechanism is ionic in nature. The need to often use molar excesses of the expensive T.dbd.O ketone over ester co-reactants in an attempt to favor the mixed coupling product, while at the same time obtaining low yields at best, makes this approach suitable only for small scale preparations. Furthermore, the well-known capricious nature of the reaction, the large amounts of TiCl.sub.3 /LiAlH.sub.4 required to effect the coupling, and the formation of by-products which are difficult to separate from the desired enol ethers also limit the commercial utility of the process. In addition, certain useful meta-substituted starting materials such as: ##STR7## cannot be used with the McMurry reagents as such substituent groups would be reduced, hydrolysed, or would take part in reductive coupling with T.dbd.O. Thus, the double bond cannot be introduced regiospecifically in every case.
Enol ethers have also been prepared by Peterson or Wittig reactions of alkoxymethylenesilanes or phosphoranes with aldehydes or ketones in basic media [Magnus, P., et al., Organometallics, 1:553 (1982); Wynberg, H. and Meijer, E.W., Tetrahedron Lett., 41:3997 (1979)]. Bronstein, 1986, above, describes the synthesis of an olefin of formula (II) above using a Wittig reaction of a phosphonium ylide with a T.dbd.O ketone. A major advantage of the Wittig reaction is that it is an ionic reaction, where the double bond can be introduced regiospecifically in almost every case.
One problem with the Wittig reaction, however, is that the product alkene is difficult to separate from the phosphine oxide by-product because of the similar solubility characteristics of these compounds. Another problem is that the initially-produced phosphonium ylides can be made only from relatively expensive phosphine starting materials [Walker, B.J., in Cadogan, J.I.G., ed., "Organophosohorus Reagents in Organic Synthesis", Academic Press, N.Y. (1978), pp. 155-205]. Also, as phosphonium ylides are relatively weakly nucleophilic, they will react only with a limited range of carbonyl compounds, and can require relatively harsh reaction conditions to do this [Gushurst, A.J., et al., J. Org. Chem., 53:3397 (1988)]. Finally, side reactions frequently occur in the Wittig reactions, which also contribute to relatively low yields [Horner, L., et al., Chem. Ber., 95:581 (1962)].
Because of the many problems attendant upon both the McMurry and Wittig reactions, particularly when used to synthesize olefinic intermediates for enzyme-cleavable 1,2-dioxetanes on a commercial scale, a more-suitable route to enol ether derivatives useful in the synthesis of stable, water-soluble, enzyme-cleavable chemiluminescent 1,2-dioxetanes was needed.